Vaccinia virus was used as the means to eradicate wild-type smallpox. The vaccine was known to have a high rate of complications, but immunization was considered desirable because of the severity of the natural disease and the concerted world- wide to eradicate the virus. Since eradication of circulating smallpox, recommendations for vaccinia virus immunization have been limited to laboratory workers who work directly with the virus, because such work may involve exposure to high titered preparations that also may have heterologous antigens introduced by recombinant technology. The rate of severe complications for primary vaccines is markedly higher than the rates of complications for most other vaccines in routine use in the US. The only available therapeutic agent for complications of vaccinia immunization at this time is vaccinia immune globulin (VIG) prepared from the plasma of vaccinia virus vaccinees, but this material is in short supply and the sources for such blood products have become more limited since the cessation of routine immunization. This project will generate a sustainable, well- characterized immunotherapeutic strategy for treatment or prevention of complications, namely human vaccinia virus neutralizing antibodies for passive administration. The research design is novel in that is uses a new transgenic mouse that expresses a human antibody repertoire (Xenomouse) to derive human monoclonal antibodies. In this model, multiple frequent vaccinations can be performed in order to ensure a high frequency of virus-specific B cells for generation of hybridomas. In the Xenomouse, B cells and T cells are of murine origin thus allowing efficient generation of high affinity antibodies during secondary responses, but the antibodies produced are fully human. Human antibodies are preferable to reenigineered or unaltered murine antibodies because the safety and efficacy of rodent antibody therapy in humans may be affected by a vigorous human-anti-mouse- antibody response. The methods to be used involve B cell hybridoma generation or direct cloning of antibody genes from B cell clones. The proteins to which the neutralizing antibodies are directed will be determined. This work will allow the logical selection of a subset of the antibodies for use in a therapeutic cocktail that differ in their specificity, so as to prevent the emergence of virus escape mutants during clinical use. In summary, this work will yield a safe and effective immunotherapeutic strategy for treatment of vaccinia complications.